Protein phosphatase 2A (PP2A) is a ubiquitous member of the protein serine/threonine phosphatase gene family functions in fundamental cellular processes including metabolism and the cell cycle. PP2A is composed of core complex containing the catalytic subunit (C) and the scaffold subunit (A), which interacts with a wide variety of regulatory subunits and interacting proteins that target the AC core dimer to specific substrates and intracellular locations. The assembly of many different types of heterotrimeric holoenzymes accounts for the ability of PP2A to regulate a wide range of biological processes. Important targets for PP2A include the neuronal microtubule cytoskeleton, the Cdc6 protein involved in DNA replication, and components of the Wnt signaling pathway. One overall goal of this project is to determine the molecular basis for the actions of PP2A in these processes. Another major goal is to identify novel functions of PP2A by identifying new phosphoprotein targets for the enzyme. The project has four specific aims. 1. Aim 1 will complete the analysis of transgenic mice in which targeting of PP2A in the brain is disrupted by expression of SV4O small tumor antigen. The experiments will test the hypothesis that forms of PP2A targeted to microtubules play an important role in regulating the phosphorylation of the neuron-specific microtubule-associated protein tau. The experiments will also test whether increased phosphorylation of tau can lead to neuronal degeneration. 2. Aim 2 will determine the role of the PR48 subunit in targeting PP2A to the Cdc6 protein. The experiments will test the hypothesis that the interaction of PP2A with Cdc6 is required to maintain Cdc6 in a dephosphorylated state during the GI phase of the cell cycle. Experiment in this aim will also determine the role of calcium in regulating Cdc6 dephosphorylation. 3. Aim 3 is designed to identify novel PP2A substrates and functions using a proteomic approach. Individual PP2A subunits will be knocked out using RNAi. The consequences of the loss of these subunits on cellular signaling will be examined by two-dimensional electrophoresis, identification of proteins by mass spectrometry, and DNA microarrays. 4. Aim 4 will test whether the ABeta subunit of PP2A is a tumor suppressor protein. Wild type Abeta protein and forms of the protein containing mutations identified in lung and colon tumors will be tested for their abilities to downregulate the Wnt signaling pathway and to reverse the tumorigenic potential of lung cancer cell lines